Glioblastoma multiforme (GBM), the most common malignant central nervous system neoplasm, is a uniformly fatal disease. Immunotherapy as a treatment for GBM has garnered increasing interest due to its potential to generate a long-lasting, highly specific anti-tumor response. However, clinical efficacy to date has been modest, in part due to tumor-induced immunosuppression. Effective immunotherapy relies on induction of tumor-specific cytotoxic T lymphocytes (CTLs) that attack the patient's tumor. However, CTL anti-tumor activity can be reduced through interactions with programmed death ligand 1 (PD-L1) present on the surface of GBM cells, tumor associated macrophages (TAMs), and circulating monocytes. As a consequence, GBM patients experience immunosuppression locally in the tumor microenvironment, and systemically in the peripheral circulation. The long-term goal of the proposed work is to improve GBM immunotherapy through the identification, characterization, and modulation of GBM derived soluble factors (GDSFs) that contribute to immunosuppression, especially through induction of PD-L1 expression in the myeloid cell population. Preliminary evidence indicates that (1) elevated peripheral monocyte PD-L1 expression predicts worse outcomes in GBM patients treated with immunotherapy, that (2) one GDSF, activated leukocyte cell adhesion molecule (ALCAM), promotes PD-L1 expression in monocytes, and (3) increased soluble ALCAM (sALCAM) is present in the plasma of patients whose tumors induce PD-L1 in monocytes. Given these relationships, it is hypothesized that GBM expression of ALCAM plays a role in the induction of PD-L1, both locally and systemically. Furthermore, inhibition of ALCAM itself, and/or ALCAM-induced signaling pathways that stimulate PD-L1 expression, is hypothesized to reduce immunosuppression by decreasing PD-L1 expression in myeloid cells. In the first aim of the proposal, we will investigate the relationship between GBM patient ALCAM levels and PD-L1 expression in circulating monocytes as well as TAMs. Moreover, we will explore how these variables affect patient survival and functional immunosuppression. The second aim will focus on identifying key signaling pathways involved in ALCAM-induced monocyte PD-L1 induction. Through these analyses, it will be possible to identify signaling mediators that can be targeted to reduce GBM-induced, monocyte-mediated immunosuppression. The third proposed aim will evaluate the role of ALCAM on anti-tumor immune responses, tumor growth, and overall survival in a murine GBM model. Through modulation of tumor cell ALCAM expression, we hope to provide preclinical evidence for the benefit of targeting ALCAM to reduce immunosuppression and promote survival. Through these experiments, we hope to gain an understanding of the mechanisms driving immunosuppression, which, in turn, can promote the development of novel strategies to increase the efficacy of immunotherapeutic approaches for treating GBM.